Infectious agents as disruptors of calcium homeostasis and possible
implications for autism

One proposed etiology for autism is viral infection very early
in development. Various viruses and other infectious pathogens
have been implicated as etiological factors, including several
members of herpes family, rubella, influenza, measles and mumps
viruses. In addition, individual cases of prenatal syphilis
and toxoplasmosis each have been reported in literature [15804954,
222139,
4947755].

The best association to date have been made between perinatal
cytomegalovirus and rubella viruses
and autism. Athough detailed large scale follow-up studies have
yet to be carried out, numerous case reports describe perinatal
cytomegalovirus infection in association with development of
autism [12959425,
6315673,
6086566].
It is estimated that up to 15% of asymptomatic congenital infections
by CMV in the neonatal period develop persistent problems, which
often involve neurological disorders and which typically appear
after a period of time, thus making diagnosis difficult or impossible
[15973639].
With regards to rubella virus, one follow-up report on psychiatric
and behavioral consequences of documented congenital infection
recorded significantly high rates of mental retardation and
behavioural pathology, including autism [702254].

A recent study found polyomavirus infection in postmortem autistic
brains BKV, JCV, and SV40 viruses were significantly more frequent
among autistic patients compared to controls [20345322].

Also worth
noting is the finding that a large subset of subjects with autism
shows evidence of bacterial and/or viral infections not present
in age-matched controls [17265454],
as well as a reduced response to vaccine antigens, especially
Bortedella pertussis [link]. In
additon, there appears to be a correlation between of virus
serology and brain autoantibodies in autism (see Immunity/Inflammation).

Viruses

Animal models in which early viral infection results in behavioral
changes later in life include the influenza virus model in pregnant
mice and the Borna disease virus model in newborn Lewis rats.
Neonatal Borna disease virus (BDV) infection
of the rat brain is thought to be a useful model for studying
the pathogenesis of neurodevelopmental abnormalities in humans,
as it produces neurodevelopmental damage in rodents that is
similar to some pathological and clinical features of autism
and schizophrenia. Amongst other things neonatal BDV infection
profoundly affects social behaviors in adult rats [16860408].
Another significant finding is that the same virus was found
to produce differential neuroanatomical and behavioral abnormalities
in genetically different inbred rat strains [12106670].

Of note is that neonatal rats infected with BDV do not mount
an aggressive response to the virus like their adult counterparts,
but instead develop a persistent infection with
less obvious clinical symptoms. Postmortem
examinations of the rats’ brains reveal significant loss
of Purkinje and dentate gyrus granule cells, as well as degeneration
of the parts of hippocampus [10321982].
Similarly, selective neuronal damage was observed in BDV replication
in hippocampal slice cultures from rats, where damage and death
of granular cells was accompanied by reduced density of mossy
fiber axons [16140749].

BDV infection in rats is also associated with elevated
expression of cytokine and chemokine mRNA, as well
as abnormal levels of neurotransmitters, including
increased tissue concentration of serotonin in parts of brain
[15385249].
Significan abnormalities in brain levels of metallothioneins
and zinc metabolism have also been observed, leading to suggestions
that such disturbances may contribute to neurodevelopmental
damage in perinatal viral infection [16612977]
(see below).

Prenatal influenza viral infection in mice
leads to development of abnormal emotional and cognitive functions
and behaviours, including deficits in social interaction, in
adult offspring [12064515,
12514227].
The infection causes differential expression of genes in brains
of the infected mice [15906383]
as well as atrophy of pyramidal neuorons and increased brain
size in adulthood [12064515].
These findings were confirmed by another study, where it was
noted that the alterations in gene expression
became apparent only after a latency period,
an observation which may be of significance in autism [15973158].

Upregulated expression of glial fibrillary acidic protein (GFAP),
an important marker of gliosis, neuron migration, and reactive
injury, has been recorded [12140787].
Of note is that significant elevations in levels of GFAP, as
well as autoantibodies to the protein, have been observed in
autism through plasma, cerebrospinal fluid and postmortem brain
examinations [16147953,
9308986,
8353169].
In addition, reduced levels of Reelin have
been observed in mouse offspring, implicating the involvement
of this protein in neuronal migration abnormalities in mouse
brains following viral infection [10208446]
(see Brain re Reelin abnormalities
in autism).

Developmental abnormalities in paediatric Human Immonodeficiency
Virus (HIV) infections are of great relevance for establishing
mechanisms that lead to autism. Children that are pre- or postnatally
infected with the HIV subsequently developed neurological symptoms,
including impairments or declines in cognitive, language, socio-emotional
and motor development. In some cases regression/deterioration
of previously acquired skills can happen several years after
the original infection, the longest reported being in a child
of 5 years of age, who previously exhibited age-appropriate
behaviours and development of all areas of concern [7682171,
7770287,
8935240].
Most importantly, improvements in neurological distrurbances
are often noted following antiviral or immune-modulation therapies.
These improvements include diminishing
of autistic symptoms and recovery, sometimes complete, of language
and cognitive abilities, normalised play and social behaviour,
and improvements in motor skills, muscle tone and visuo-spatial
functions [2493377,
9493492]
further reading: [link]

The cental mechanism behind the neurodevelopmental decline and
behavioural problems in paediatric HIV infection is thought
to be the neuronal injury caused by excessive increases
of intracellular calcium induced by HIV viral proteins
[16553776,
7847672].
In addition to HIV, numerous other viruses, including many that
have been implicated in autism, code for proteins that are capable
of inducing similar perturbances in calcium homeostasis in neurons
and other types of cells. Calcium plays an important role in
replication cycles and pathogenesis of many viral diseases and
viruses have been proposed to influence calcium homeostasis
of host cells through several different pathways.

An estimated third of the adults and half of the children with
acquired immunodeficiency syndrome (AIDS) induced by HIV exibit
neurological disturbances. This is usually refered to as AIDS-induced
dementia, neuro-AIDS or HIV encephalitis, and the symptoms include
language and attention problems, lethargy, motor and sensory
dysfunction and abnormal reflexes. Affected children exibit
many autistic symptoms, including language and socio-behavioral
abnormalities (see above). Some of the pathologies reported
in the brain of patients with AIDS are neuronal injury and loss,
including myelin loss and axonal damage, and microglial activation.

Dysregulation of calcium homeostasis underlies neurological
disturbances in neuro-AIDS. The virus seems to enter the brain
in the early stages of infection and arising pathological processes
in the brain and CNS are only in part affected by HIV-dependent
processes in the periphery. Although the virus does not seem
to directly infect the neurons, two of its identified viral
proteins are neuroxic - the viral coat protein gp120 and the
transcription regulator Tat. Both of these proteins can induce
apoptosis of cultured neurons and can render neurons vulnerable
to excitotoxicity and oxidative stress.
Both gp120 and Tat disrupt neuronal calcium homeostasis by perturbing
calcium-regulating systems in the plasma membrane and endoplasmic
reticulum. By altering voltage-dependent calcium channels, glutamate
receptor channels, and membrane transporters, these proteins
promote excessive calcium entry, production of free radicals
and mitochondrial dysfunction [12394783]
(see Oxidative_Stress and
Mitochondrial issues in autism).
These effects are at least partly mediated via their effect
on chemokine receptors, which are widely expressed
on neurons and some of which are able to activate the calcium
and cAMP-dependent transcription factor CREB
(see Brain) [9826729].
In addition, glial cells were also found to contribute to chemokine
upregulation in the CNS and so to contribute to neurological
damage [15163738,
11744246].
(see chapter below re chemokine receptors on neurons and glial
cells).

Of
particular relevance to autism could be the observation of the
ability of retroviruses to cause neurological damage in the
absence of lympocyte infiltration, primarily through upregulation
of chemokine MCP-1 [15163738].

Decreased plasma ratio of tryptophan observed in HIV infected
subjects has also been sugested to have possible implications
for development of neuro-AIDS. Low levels of tryptophan have
been proposed to cause a decrease in serotonin synthesis
in the brain, which is believed to impair functioning of some
types of neurons [9026369].
Brain serotonin level is suspected to play an important role
in developing brain and impairments of serotonin metabolism
have been implicated in autism. Reduced brain levels of tryptophan
and/or serotonin and its receptors have been recorded in other
viral infections. Several in vitro studies have noted the inhibitory
effects of serotonin and its receptors on the function of VGCC
and neuronal migration during development [11976386]
(see also Brain and Maternal_Factors).

For
further details and references on retrovirus-induced autism
and physiopathological parallels to idiopatic autism go to HIV_and_autism
page.

Various calcium antagonists, acting on both voltage channels
and glutamate operated NMDA receptors, are effective in vitro
in reducing neuronal damage induced by HIV proteins. Reductions
in glutamate levels also protected neurons from gp120-induced
injury, suggesting that they act synergestically and that both
are necessary for neuronal cell death [1656845].
Calcium channel blocker nimodipine has shown promise for HIV-induced
neurological disorders in a Phase I/II clinical trial [9674806].

In addition to HIV, Maedi Visna virus, another lentil virus,
often causes a variety of neurological symptoms in infected
sheep. This effect once again is thought to be mediated via
neurotoxic effects of its Tat protein and could be attenuated
in vitro by application of calcium channel antagonists [8552302].

The effect of HIV proteins on chemokine receptors and modulation
of calcium fluxes in immune cells is though to play an important
role in HIV-induced immune dysfunction (see Immune/Inflammation).
Similar mechanisms are thought to lie behind HIV disturbances
of gastrointestinal function (see Gastrointestinal).

Several viruses from the herpes family have
been implicated in the etiology of autism (see above). Some
of the herpesviruses are neurotropic and are involved in the
pathogenesis of neurological symptoms in infants. Cytomegalovirus
(CMV) infection is the most frequent congenital infection in
humans and can cause permanent damage, often following initial
asymptomatic period. The most frequent symptoms are mental retardation,
hearing loss, microcephaly as well as hydrocephalus, periventricular
calcification, neuromuscular disorders and chorioretinitis or
optic atrophy [6159568].

In vitro CMV was found to prevent induction of differentiation
of human neural precursor cells into neurons. The virus arrested
cell growth and induced apoptosis in infected cell cultures
and this effect of CMV was proposed as a possible explanation
for the abnormalities in brain development associated with congenital
infection [16940505].
A study looking at intracellular calcium responses to CMV infection
noted excessive influx of calcium shortly after the infection
of cultured human fibroblasts, with substantial involvement
of intracellular stores in addition to voltage gated plasma
channels. Application of calcium channel blockers inhibited
the rise in intracellular calcium levels. Authors of the study
suggested that the observed effects of the virus on calcium
metabolism may be related not only to the development of CMV
pathology, but also to viral replication itself, since in other
studies cyclic nucleotide modulators and calcium influx blockers
were found to inhibit the replication of CMV [3029971].

CMV-induced rise of intracellular calcium is thought to be behind
the ability of this virus to interfere in cellular differentiation
pathways, including inhibition of macrophage differentiation,
thus contributing to CMV immunosupressive effects [15470031].

Significant abnormalities in calcium metabolism of influenza
virus-infected cultured cells are brought on by enhancement
of permeability of membrane calcium channels and also by mobilizing
calcium from intercellular stores [3135328,
6626708].
Similar to CMV, influenza viral replication could be inhibited
in vitro by a calcium channel blocker, as well as by a drug
which binds to calmodulin and so inhibits calcium/calmodulin
intracellular pathways that are necessary for virus assembly
[6743023,
6808973].One
of the proteins encoded by coxsackie virus, protein 2B, increases
membrane permeability and release of calcium stored in endoplasmic
reticulum, and in this way disrupts cellular calcium homeostasis,
eventually causing membrane lesions that allow release of virus
progeny [9218794,
12903773].
Similar effects have been noted following infection of cultured
human fibroblasts by poliovirus, another enterovirus
from Picornavirus family [7609085].
(see also Immune/Inflammation)

Coxsackievirus B3, a strain capable of inducing
myocarditis, has been observed to increase
calcium inflow through LTCC in cardiomyocytes, an affect which
could be inhibited in vitro by aplication of calcium antagonist
taurine [10375733].
Calcium blocker verapamil had similar effects in a murine model
of virally-induced inflammation of heart muscle. Pretreatment
with the drug before and during the infection, as well as administration
of the drug several days after the infection significantly reduced
the microvascular changes and myocardial necrosis, fibrosis,
and calcification leading to cardiomyopathy [1331179].

Of particular relevance to autism could be the observation concerning
the role of autoantibodies to beta adrenoceptors,
since these g-protein linked receptors are expressed in the
brain as well as the heart and their activation is linked to
cAMP and membrane calcium channels. A study looking at the effects
of autoantibodies against beta(1)-adrenoceptor in hepatitis
virus myocarditis found that they were capable of inducing arrhythmias
and/or impairment of heart mucle, an effect that is most likely
mediated by LTCC [15069720].

In a similar manner cardiac abnormalities expressed in congenital
heart block in newborns born to mothers with autoimmune
disease are caused by maternal autoantibody-mediated disturbance
of LTCC function [11257091]
(see Maternal_Factors).

Mouse hepatitis virus, a murine coronavirus,
induced rapid and transient increases in intercellelar calcium
via LTCC in a small percentage of cells infected in vitro. It
was concluded that the cells that responded with enhanced calcium
signals were those that had been infected with multiple viruses
and undergone rapid viral replication. Furthermore, several
calcium channel blockers and the calcium chelator EGTA inhibited
virus infection in this model [9417866]. Several
viral proteins Human hepatitis C virus, appear to contribute
to reactive oxygen species (ROS) generation by mechanisms that
involve dysregulated calcium metabolism, including calcium uptake
by mitochondria [16958669]
(see also Mitochondria).

Mumps virus, which causes a persistent non-lytic
infection, was shown to alter the function of membrane and intercellular
calcium channels in cultured neuronal and glial cells in several
studies. It has been suggested that this occurence may reflect
a disturbed glia-nerve cell interactions [1647243, 1647243].
Although a reduction in the influx of calcium ions during early
stages of infection by mumps virus was recorded, neuronal degradation
was almost completely inhibited by nifedipine in one study and
by dantrolene, a drug which inhibits release of calcium from
intracellular stores, in another [1662994,
9372458].

Apart from its better-known immunosupressive role, it is believed
that in rare cases measles virus may cause
progressive neurodegenerative disease [14527283].
Inhibitory effect of the calcium antagonist, verapamil, on measles
viral replication in cultured cells has been observed [8454440].
However another study concluded that this effect was probably
not related to verapamil block of extracellular calcium entry.
It is suggested that, similar to the abovementioned effects
of calcium modulators on influenza virus, this is due to drug
interferance in the calcium/calmodulin intracellular
pathways that are necessary for virus assembly [16054245].
On the other hand the immunosupressive effect of measles virus
likely caused by one of its protein binding to nucleoprotein
receptor on cell surface, which triggers sustained calcium influx
and inhibits spontaneous cell proliferation [14557619].

Rotavirus tends to effect gastrointestinal
epithelial cells, causing acute gastroenteritis and occasionally
lactose intolerance (see Gastrointestinal).
Growing line of evidence in recent years is pointing to the
ability of the virus to leave gastrointestinal tract and cause
a broad range of systemic diseases in a number of different
organs, including the CNS. Several case studies describe patients
who developed seizures and other neurological problems
following rotavirus infection. In one particula case diffuse
inflammation of the brain was detected by MRI. Neurological
symptoms in the patient included screaming fits and seizures,
loss of language and social interaction,
and reduction in muscle tone. These symptoms
persisted in long after the disappearance of gastrointestinal
problems [11731961,
12454200].

Animal studies detected replicating rotavirus in infected mice
in, amongst other places, macrophages in the lungs and blood
vessels, indicating a possible mechanism of its dissemination
throughout the body. Interestingly, viral spreading and replication
was noted even in the absence of diarrhea. Of note is that in
the animals that were infected past a certain stage of development
the virus seemed not to infect the brain, as was the case in
younger animals. It was concluded that dissemination of rotavirus
to the brain in mice is age restricted [16641274].

The observation of the interaction of rotavirus protein NSP4
with membrane protein caveolin-1 during viral
infection [16501093]
could prove to be of some significance in autism and/or epilepsy,
since caveolin-1 is expressed by neuronal cells and seems to
exert significant effect on functioning on several ion channels,
including VGCC, and on cellular excitability [16040758]
(see Membrane_Metabolism).
In additon, caveolin-1 seems to play an important role in some
inflammatory pathways and in the maintainance of Blood
Brain Barrier (BBB) – its loss is proposed to
be a critical step in MCP-1-induced modulation of brain microvascular
endothelial cells junctional protein expression and integrity
[17023578].
Similarly, its downregulation in murine macrophages was shown
to increase LPS-induced proinflammatory cytokine TNF-alpha and
IL-6 production, but decrease anti-inflammatory cytokine IL-10
production [16357362].

Bacteria and other infectious agents

Bordetella pertussis is a gram-negative bacteria,
the causative agent of pertussis (whooping cough). Its toxin
interferes with intracellular communication by affecting the
functioning of regulatory G-proteins and preventing them from
interacting with cell membrane receptors. The close involvement
of G-protein activation in regulation of opening times of voltage
gated calcium channels has been observed in many types of cells
including neurons [8382734,
2560167],
insulin-secreting pancreatic betta cells and catecholamine-releasing
adrenal chromaffin cells [15488596,
1714959]
(see GI, Neurotransmitters
and Hormones).

In animal studies the changes that were observed with regards
to permeability of BBB and cellular calcium
overload were thought to play a central role in the pathogenesis
of infectious brain edema induced by injection
of pertussis bacilli in rat brain. Treatment with nimodipine
was neuroprotective – it dramatically reduced the damage
of brain edema by inhibiting the excess of calcium influx and
reducing the permeability of BBB (see BBB).
Pretreatment of neurons by MK-801, a NMDA channel antagonist
was observed to inhibit delayed calcium influx [12659709,
9812737].
Similarly to Borna Disease Virus infection of neonatal mice
(see above), the reaction to pertussis toxin/vaccine and occurrence
of toxin-induced encephalitis appears to be mouse-strain dependant
[6933900].
In addition, it was found that mice that overexpress monocyte
chemoattractant protein-1 (MCP-1) at high levels show greater
vulnerability to pertuss toxin and that the disruption of CC
chemokine receptor 2 (CCR2) abolished both CNS inflammation
and encephalopathy in this mouse model, identifying the central
role of CCR2 in pertussis-induced encephalopathy
[12486156]
(see below on role of chemokine receptors in neurological disorders).

A significant association was shown between neurological disorders
and pertussis toxin in humans [6786580,
9755273].
Residual levels of active pertussis toxin and endotoxin are
likely to be a major contributors to the reactogenicity of whole
cell pertussis vaccines, and new guideliness and limits have
been established by WHO for active pertussis toxin in acellular
pertussis vaccine [link].
In addition, it has also been proposed that permeability changes
in the cerebral vessels may be involved in the evolution of
the encephalopathy attributed to the use of the vaccine [12780,
12503649].

Various other types of bacteria and their toxins interfere with
calcium homeostasis of the host cell. Bacterial lipopolysaccharide
(LPS) activation of mouse microglia in vitro leads to elevated
basal calcium along with attenuated calcium signaling in response
to stimulation, translating into reduced ability of activated
microglia to respond to an external stimulus. In addition, the
rise in calcium levels underlie characteristic features of microglial
activation, such as release of nitric oxide (NO) and several
cytokines and chemokines. In the absence of available calcium
the LPS-stimulated secretion of cytokines and NO is greatly
reduced.

One of the proposed ways of bacteria-induced rises in basal
calcium is through formation of toxin-created calcium channels
on cell membrane required for bacterial cell invasion, although
more recent line of evidence suggests direct modulation of existing
LTCC channels and their function [3917612,
6510944,
16732377,
12736823].
Addition of calcium channel blockers, calmodulin antagonists
and/or chelation of extracellular calcium significantly reduce
bacterial entry into host cells in vitro [12761148,
8382566,
16151220].
This same effect was also noted in invasion of host cells by
Toxoplasma gondii [9309395,
15591836],
a parasitic agent capable of passing on to humans and causing
low grade encaphalitis followed by neurological and behavioural
modifications [16000166].

Activation of various chemokine receptors expressed
on cell surface by bacterial agents is proposed as another mechanism
behind bacteria-induced rises in cell calcium levels (see below).
Apart from the actions of their LPS, other bacterial membrane
components are also known to act on various chemokine and toll-like
receptors and influence their expression in different cell types,
with CCR2, CCR5, TLR2 all being implicated [10559223,
14733721,
15885315].
TLR2 for example is essential for the recognition of peptidoglycan
and lipoprotein/lipopeptides that are present
in cell walls of a variety of micro-organisms, and rodent studies
show that mice lacking this receptor do not exibit inflammatory
response when exposed to these agents [12697090].

Encaphalitis caused by mycoplasma pneumoniae,
a lung-infecting bacteria capable of causing neurological manifestation,
is characterised by elevated levels of several cytokines and
beta-chemokines in the lungs and the CNS [10441731,
16087054].
Changes in the intracellular calcium levels, most probably mediated
through activation of toll-like receptors TLR2 and -6, are though
to underlie the systemic inflammation and pathologenesis of
mycoplasma genus of bacteria [16154916,
15312143].
In addition to the involvement of G-protein coupled receptors
and plasma calcium channels, subsequent release of calcium ions
from endoplasmic reticulum has also been observed [11953388].

Lipoprotein
found on Treponema pallidum, causative agent of syphilis
and implicated as one of the possible causative or contributing
agent in autism (see above), induces CCR5 on human monocytes
and enhances their susceptibility to infection by HIV [10608777].
Absence of the chemokine receptor CXCR2 results in reduced inflammation
induced by Borrelia burgdorferi, another spirochete
bacteria and the causative agent in Lyme disease and neuroborreliosis
[15760769].
Similarly, infection of nul CXCR2 mice resulted in a significant
decrease in susceptibility to development of experimental Lyme
arthritis, suggesting that chemokine-mediated recruitment of
neutrophils into the infected joint is a key requirement for
the development of this condition [12847259].

Prion encelopathies are caracterised by neurotoxicity,
neurodegeneration and microglial activation induced by mutated
forms of the prion protein and the amyloid beta precursor protein
(Abeta), and are thought to be mediated through their effects
on calcium conductances through L-type calcium channels [9914452,
14622132,
10817932].
The so called channel hypothesis of Alzheimer's disease proposes
that Abeta proteins accumulate in the brain and damage neurons
by forming ion channels [12128087].
This hypothesis is further supported by neuroprotective effects
shown by some calcium channel blockers in experimental Abeta
neurotoxicity [17051460].
In addition, the involvement of presenilin enzymes in Azheimer’s
disease is hypothesised to be unrelated to their role in formation
of amyloid peptides, but more likely linked to ability of mutated
forms of presenilins to directly disturb cellular calcium homeostasis,
leading to excessive levels of intracellular calcium, formation
of ROS and cellular death [9614221]
(see Related_Conditions).

Chemokine receptors and calcium homeostasis in the brain

Chemokines belong to a family of chemotactic cytokines that
direct the migration of immune cells towards sites of inflammation.
They mediate their biological effects by binding to chemokine
receptors on the cell surface. In adition to cytokines and chemokines,
various other agents, like viral and bacterial proteins are
capable of binding to these receptors and influencing cellular
metabolism through this mechanism (see above). Since both chemokines
and their receptors as well as various infectious agents have
been implicated in the pathophysiology of a number of autoinflammatory
diseases, including those involving inflammatory processes in
the brain and CNS, chemokine receptor antagonists have often
been suggested as potential treatment agents.

Chemokine receptors and their ligands are thought to have regulatory
functions in the normal nervous system, where they participate
in cell communication and may play a role in the control of
cerebellar neuron survival and development. Since many of these
receptors are widely expressed by various cells belonging to
both the immune and the nervous systems, they have been suggested
to serve as a ‘bridge’ between the two. This is
further supported by the observed crosstalk between chemokine
and neuropeptide receptors, such as opioid, vasoactive intestinal
peptide, or adenosine receptors, whereas activation of one often
has inhibitory effect on the other [16204635]
(see also Related_Channels). In addition, activation and/or
inhibition of these G-protein coupled receptors has a direct
effect on functioning of membrane calcium channels and cellular
calcium homeostasis.

In the CNS,
activation of neuronal chemokine receptors
by their ligands induces calcium transients and activates calcium
cAMP-dependent gene transcription factor CREB (see
Brain re its role in neuronal
gene expression). Of note is that these effects were in some
cases observed to be pertussis toxin-insensitive, indicating
possible involvement of pathways that are not mediated by G-proteins
[11958818].
Increases in calcium levels have also been observed in cultured
embryonic and adult neural progenitor cells and embryonic
neurons, with neurospheres grown from young mice exhibiting
significantly larger increases in calcium following chemokine
treatment than did neurospheres grown from older mice. It was
suggested that chemokines may play an important role in control
of progenitor cell migration in embryonic and adult brain [15048927,
11567789,
15465598]
(see Brain re expression
and function of calcium channels during brain development).
In addition, the calcium/CREB-mediated upregulation of chemokine
receptors mRNA has been observed in T lympocytes,
whereby activation of for example CXCR4 by viral proteins leads
to increased viral infectivity [11874984,
14563373]
(see Immune/Inflammation
re proposed involvement of cellular calcium loading in reduced
T-cells responsiveness in autism).

Of special significance to developmental disorders could be
the possible role played by chemokine receptor CCR2,
as its involvement in neuronal communication and in particular
cholinergic and dopaminergic neurotransmission
has been observed. Its preferential ligand, chemokine MCP-1/CCL2
was shown to induce calcium transients in primary cultured neurons
from various rat brain regions including the cortex, hippocampus,
hypothalamus, and mesencephalon [16196033].
The near-universal induction of MCP-1 and CCL2 after neurological
insult suggests that this chemokine plays a central role in
the physiology of neuroinflammation – in animal studies
chronic overexpression of MCP-1 in the CNS causes delayed encephalopathy
and impaired microglial function in mice, and possible dysfunction
of BBB has also been implicated. This experimental pertussis
toxin-induced reversible mouse encephalopathy was completely
dependant on MCP-1 overexpression, and mice lacking the MCP-1
gene showed absolute resistance [15857890].
Of note is that levels of MCP-1 were found in a postmortem examination
to be significantly raised in brains and CSF (12-fold rise)
of subject with autism compared to controls (see Immunity/Inflammation).

Similarly, overactivation of other chemokine receptors, as well
as toll-like receptors, have been implicated in CNS inflammatory
diseases [17151286,
15557205].
The presence of fractaline receptor CX3CR1 signalling has on
the other hand recently been proposed as having protective effects
against microglial neurotoxicity [16732273]

Apart from
neurons, chemokine receptors are also widely expressed on brain
glial cells, where their function modulates
intracellular calcium signalling and calcium-regulated gene
expression [15850656,
16547971,
12112372].
The abovementioned interactions between chemokine and opiod
receptors and downstream effects on calcium-mediated pathways
are thought to underlie the observed effects of opiod
treatment and antagonism on viral infection and outcomes
[15893700].
Cultured astrocytes simultaneously treated with opiates and
viral protein Tat exibit exaggerated increases in chemokine
release, including MCP-1, RANTES and IL-6, an effect that is
mediated via the modulation of MAPK and CREB signaling pathways,
and that can be prevented by applying a mu-opioid receptor antagonist
[15630704,
15893700,
17162664]
(see also Related_Receptors).

Genetic polymorphisms as predictors of inflammatory outcomes
and chemokine receptors as treatment targets

Several chemokine receptor gene variants have been demonstrated
to have detrimental effect on HIV infection and progression
to AIDS, and various CCR5 genotypes in children determine both
speed of disease progression as well as the degree of
neurological impairment [14624371,
9696841].

Similar chemokine receptor-related genetic polymorphisms play
a role in vulnerability of gastrointestinal tract
to infectious agents, and chemokines and their receptors are
thought to be viable therapeutic targets for the treatment of
inflammatory bowel disease. A good example is that genetic deficiency
in the chemokine receptor CCR1 was shown to protects against
acute Clostridium difficile toxin A enteritis in mice - the
toxin induced in all mice a significant increase in ileal fluid
accumulation, epithelial damage, and neutrophil infiltration,
but with all parameters being significantly lower in CCR1 and
MIP-1alpha knockout mice [11875005].

Targeting CCR5 and its ligands has been suggested as a possible
immune-modulating strategy since it appears
to be involved in recruitment of immature MHC class II dendritic
cells to the site of inflammation [15790880]
(see also Immunity/Inflammation).
In the case of HIV infection, various agents capable of modulating
expression of multiple chemokine receptors are currently being
developed as novel antiviral strategies [12936978,
12936978,
7000001].

The blockade of CCR2, CCR5 and CXCR3 by a novel chemokine antagonist
TAK-779 prevents experimental colitis in murine
studies by inhibiting the recruitment of inflammatory cells
into the mucosa [16000328].
TAK-779 is currently in clinical trial for human use approval
and is expected to be used as treatment agent in HIV infection and
possibly Inflammatory Bowel Disease (see Gastrointestinal
for inflammation and other gastrointestinal issues in autism).

Other issues for consideration

Observations that a virus can act both as an immunosupressor
and as a stressor, capable of activating
the hypothalamic-pituitary-adrenal axis and increasing brain
concentrations of tryptophan and norepinephrine catabolite should
be given some consideration in the context of brain development [2756050, 3509812] (see Hormones
and Maternal_Serotonin).

A series of murine and studies pointing to links between
viral infections and absorbtion and tissue/organ distribution
of environmental toxins may be of great relevance.
It has been observed for example that the intestinal absorption
of cadmium increases during a common viral infection - in the
infected animals the absorption of this metal was increased
by 70% at low doses and was tripled at high doses compared to
non-infected animals. The increased absorption also enhanced
the accumulation of cadmium in all studied organs [9630849].
Redistribution of trace elements already present in the body
has been noted, with brain levels of mercury increasing
twofold during viral infection [16327074].
Mercury was shown to change virally-induced myocarditis in a
direction compatible with the development of chronic disease
and allow increased persistence of virus, indicating that heavy
metals may interact and adversely affect viral replication and
development of inflammatory disease. Of note is that in the
inflammatory lesions of mice exposed to mercury, the myocardial
contents of calcium, manganese, and iron were significantly
increased compared to controls [11314973].
The observed changes in the patterns of accumulation, excretion
and toxicity of the environmental pollutants during common virus
infection has been proposed to be due to down-regulation
of detoxifying processes in favour of acute-phase protein
synthesis in the host in response to infection [11846173].

With regards to latent versus active viral infection, various
factors are known to be capable of reactivating a latent
virus, a process which appears to be calcium-dependent
and involves elevations of cellular calcium levels [12692270].
An estimated three quarters of diagnosed autism cases seem to
fall into category of regressive autism – a sudden loss
of previously acquired language and sociobehavoural skills,
usually occuring between 18-24 months of age, following a period
of normal deveopment. The exact reason for this has been a subject
of controversy, with parents and carers reports of regression
following series of vaccination contrasted by epidemilogical
studies reporting no causal link, leading to recent proposals
to study incidence of autism in unvaccinated populations. Of
note is that vaccinations have been proposed as capable of triggering
reactivation of latent virus infections, via vaccine-induced
immunomodulation [11103441].
This mechanism of an induced immune challenge having a trigger-like
effect on latent viruses has been suggested to underlie development
of CNS autoimmune diseases [11517396].
Pregnancy has been suggested as another immune-related event
capable of triggering reactivation of latent viruses. Reactivation
of HHV-6 seems common during pregnancy, and transfer of this
herpesvirus to the fetus is estimated to occur in approximately
1% of pregnancies [10558965].

On the other
hand, latent a viral infection, notably by herpesviruses, has
also been shown to alter neuronal gene expression, including
genes involed in neurotransmittion and cellular excitability,
and latent viruses are therefore proposed to play a possible
role in etiology of chronic disease [12915567].

Finally, following recent discoveries of human chromosomal integration
of herpesviruses and their sites, the possible role in etiology
of viral integration in chromosomal abnormalities and various
human pathologies, including autism, should not be completely
discounted [15255601,
10477678,
16597897,
16341055,
4285072]
(see also Genetic_Factors).